This invention relates generally to solutions useful as diluents in the biological sciences which are commonly known as "buffers". Such solutions are capable of neutralizing both acids and bases and thereby maintain the original acidity or basicity of the solution. More particularly, the invention relates to an improved buffer useful for the dilution of immunohematological, immunological and immunochemical assay components, such as antibodies and antigens.
Immunohematological analysis of the blood is performed routinely on the red blood cells of donors and recipients in order to determine whether the red blood cells of a possible blood donor is compatible with with red blood cells an intended recipient. The standard analysis performed in these laboratories is an ABO-Rh blood typing, with the determination of other blood groups, such as Kell and Duffy, sometimes required in order to complete the analysis or so-called "match" of the red blood cells. The ABO blood group is comprised of antigens A and B, with the determination of the blood groups A, B, AB and O based on an agglutination reaction of the test sample with the anti-A, anti-B and anti-A,B antisera. The Rh, or Rhesus blood group comprises several antigens, including the antigens C, D, E, c, and e. Test samples which do not react with anti-D (anti-Rh.sub.o) are preliminarily classified as "Rh negative". These presumptive Rh negative test samples are retested with a manual antiglobulin test in which elevated incubation temperature, washing, and the use of a secondary antibody is required to confirm the "negative" result.
No direct agglutination technique, manual or automated, has demonstrated equivalent or greater sensitivity than the manual antiglobulin test with respect to detection of the D antigen. Several attempts have been made to improve the sensitivity of the anti-D reagent in order to eliminate the necessity for performing confirmatory testing. Blood grouping reagent manufacturers have added various components to their reagents to enhance the quality of the agglutination of tests performed in tubes. Thus, for example, a monoclonal IgM anti-D antibody has been included to supplement the human plasma IgG source, with the aim being to provide improved sensitivity for the determination of agglutination of the test sample with anti-D antisera. See, for example, Product insert, Ortho Diagnostics Systems, Blood Grouping Serum Anti-D (Anti-Rh.sub.o) (polyclonal and monoclonal blend). However, even the use of this reagent to improve agglutination does not eliminate the need for the use of the confirmatory testing techniques required for anti-D antisera negative determinations, or to dilute the antibody used in the determinations.
Traditional methods for immunohematological analysis which utilize microtiter methods, both manual and automated, require the use of undiluted or low dilutions of the antibody components in order to perform the immunohematological blood group analysis with specificity and sensitivity. No diluent buffer is known to be available commercially for blood grouping reagents. Products are available to add directly to the microplate to increase sensitivity, but these products only are added in conjunction with washing steps and a secondary antibody. See, for example, Product Insert, Gamma Biologicals, Micro-U Enhancement Reagent (Low Ionic) for Antibody Detection tests in Microplates, February (1986). Thus, these methods typically do not offer much of a cost savings to the user, since the user is not able to dilute the reagents required to perform the test.
Other blood group reagent manufacturers have incorporated high molecular weight additives, high protein and surfactants into their reagents to enhance aggultination reactions. Applying these reagents to microplate methodologies, however, also requires no or a very low dilution of antibody reagent in isotonic NaCl or Bovine Serum Albumin (BSA). See, for example, Product Insert, Ortho Diagnostics Systems, Blood Grouping Reagent Anti-D for Slide and Modified Tube Tests, and F. K. Widman, American Association of Blood Bank Techniques, "Serologic Techniques and Alloantibodies," pages 435-451 at 440 (1985). Further, pretreatment of the microplate, such as treatment with an anti-static gun or prewetting, previously has been required with some systems. F. K. Widman, American Association of Blood Bank Techniques, "Serologic Techniques and Alloantibodies," pages 435-451 at 439 (1985).